Source: BAYLOR COLLEGE OF MEDICINE submitted to
GENOME SEQUENCING OF STREPTOCOCCUS INIAE, AN EMERGING PATHOGEN OF AQUACULTURE
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
TERMINATED
Funding Source
NRI COMPETITIVE GRANT
Reporting Frequency
Annual
Accession No.
0205306
Grant No.
2006-35600-16569
Project No.
TEXR-2005-05191
Proposal No.
2005-05191
Multistate No.
(N/A)
Program Code
23.2
Project Start Date
Dec 1, 2005
Project End Date
Nov 30, 2009
Grant Year
2006
Project Director
Highlander, S. K.
Recipient Organization
BAYLOR COLLEGE OF MEDICINE
(N/A)
HOUSTON,TX 77030
Performing Department
(N/A)
Non Technical Summary
Aquaculture, or the managed cultivation of fish and shellfish, is a rapidly developing and expanding industry in US and foreign markets. In 1999, private aquaculture production in the US yielded 842 million pounds of product, with a value of nearly one billion dollars. Nevertheless, the US imported over 1.8 billion kilograms in 2000, having a value of 10.1 billion dollars and causing a trade deficit second only to the import of petroleum products. One of the most significant bacterial pathogens of farmed fish is Streptococcus iniae, an endemic, beta-hemolytic, non-Lancefield streptococcus. S. iniae has been shown to cause disease in a wide variety of finfish, including tilapia, trout and hybrid striped bass. During epidemic outbreaks in closed pond systems, mortality can reach 75% and losses of more than $150 million per year occur. These losses are likely to increase as production of farmed fishes expands. The goal of this project is to produce a complete, closed and finished ca. 1.8 Mb genome sequence of Streptococcus iniae strain 9117 (33% GC). Because the niche that S. iniae occupies is very different from those occupied by other streptococci, we expect to identify major differences between S. iniae and other streptococcal species. Sequencing The goal of this project is to produce a complete, closed and finished ca. 1.8 Mb genome sequence of Streptococcus iniae strain 9117 (33 percent GC).
Animal Health Component
(N/A)
Research Effort Categories
Basic
100%
Applied
(N/A)
Developmental
(N/A)
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
3113714108090%
7123713110010%
Knowledge Area
311 - Animal Diseases; 712 - Protect Food from Contamination by Pathogenic Microorganisms, Parasites, and Naturally Occurring Toxins;

Subject Of Investigation
3713 - Striped bass; 3714 - Tilapia;

Field Of Science
1080 - Genetics; 1100 - Bacteriology;
Goals / Objectives
The objective of this project is to produce a complete, closed and finished ca. 1.8 Mb genome sequence of Streptococcus iniae strain 9117.
Project Methods
Sequencing will be accomplished using a combination of random shotgun sequencing of small insert (2 kb) libraries (SIC), medium (10 kb) insert clones (MIC) and large insert (30 kb) libraries (LIC). All DNA sequencing will be performed at the Baylor College of Medicine Human Genome Sequencing Center (HGSC). Our strategy involves performing paired-end sequencing of SICs to 8-fold coverage, followed by paired-end sequencing of about 1500 MICs. Paired-end analysis will allow creation of assembled scaffolds that can further be closed into a contiguous sequence through a combination of end-sequencing of LICs and overgo hybridization to link LICs. Remaining gaps will be closed by PCR and primer walking. Sequence quality will be assessed using the phred base-calling program and only sequences with phred scores of 20 or higher will be included in assemblies. Final assembly will be verified by fingerprinting of LICs that form a contiguous tiling path on the genome. Sequence information will be made available on a website at the HGSC.

Progress 12/01/05 to 11/30/09

Outputs
OUTPUTS: We have completed the sequencing, assembly and annotation of the genome of Streptococcus iniae. The organism is 2.1 megabases in length and encodes approximately 2100 genes. A technician was hired to curate the annotation of this organism. This led to communications with other experts in the field and sharing of annotation on a website. Analyses are ongoing and a manuscript describing the findings from the sequence is in progress. PARTICIPANTS: Participants in this project include Dr. Sarah Highlander, PI, Dr. Xiang Qin, bioinformatician, and two research assistants as finishers. A technician, Lindsey Rutland, was hired to manually curate the automatically annotated genome. The sequencing was performed by the library, sequencing and finishing teams within the Human Genome Sequencing Center at Baylor College of Medicine. TARGET AUDIENCES: Target audiences include scientist interested in streptococcal pathogenesis and producers interested in methods for prevention of S. iniae disease, particularly in farmed fish. PROJECT MODIFICATIONS: A major change in sequencing strategy occurred during the project. Instead of performing high coverage Sanger sequencing, we switched to the 454 platform.

Impacts
Sequencing and annotation of the Streptococcus iniae has contributed to a change in knowledge by contributing the gene set of important pathogen of fin fish to the research community.

Publications

  • No publications reported this period


Progress 12/01/06 to 11/30/07

Outputs
OUTPUTS: During this period, the genome sequence of Streptococcus iniae was completed. The genome sequence is 2,079,160 nucleotides in length and is predicted to encode about 2000 protein-encoding open reading frames. Open reading frames have been predicted using Glimmer and GeneMark and these sequences have been compared to the NCBI NR, NCBI CDD, MEROPS protease, Sanger transporter, and PSORT-B databases to provide data to support annotation of the genome. PARTICIPANTS: Participants in this project include Dr. Sarah Highlander, PI, Dr. Xiang Qin, bioinformatician, and two research assistants as finishers. The sequencing was performed by the library and sequencing teams within the Human Genome Sequencing Center at Baylor College of Medicine. TARGET AUDIENCES: Nothing significant to report during this reporting period. PROJECT MODIFICATIONS: A major change in sequencing strategy occurred during the project. Instead of performing high coverage Sanger sequencing, we switched to the 454 platform.

Impacts
None at this time.

Publications

  • No publications reported this period


Progress 12/01/05 to 12/01/06

Outputs
Aquaculture, or the managed cultivation of fish and shellfish, is a rapidly developing and expanding industry in US and foreign markets. In 1999, private aquaculture production in the US yielded 842 million pounds of product, with a value of nearly one billion dollars. Nevertheless, the US imported over 1.8 billion kilograms in 2000, having a value of 10.1 billion dollars and causing a trade deficit second only to the import of petroleum products. One of the most significant bacterial pathogens of farmed fish is Streptococcus iniae, an endemic, beta-hemolytic, non-Lancefield streptococcus. S. iniae has been shown to cause disease in a wide variety of finfish, including tilapia, trout and hybrid striped bass. During epidemic outbreaks in closed pond systems, mortality can reach 75% and losses of more than $150 million per year occur. These losses are likely to increase as production of farmed fishes expands. The goal of this project is to produce a complete, closed and finished ca. 1.8 Mb genome sequence of Streptococcus iniae strain 9117 (33% GC). Because the niche that S. iniae occupies is very different from those occupied by other streptococci, we expect to identify major differences between S. iniae and other streptococcal species. Sequencing was accomplished using the 454 Life Sciences pyrosequencing technology. Using 454 sequences, the genome has been assembled into 199 scaffolds with an N50 contig size of 31 kb. The coverage is 28X and the genome size is estimated to be 1.96 Mb. Remaining gaps will be closed by PCR and primer walking. Once the genome is closed or nearly closed manual annotation will begin.

Impacts
With respect to intellectual merit, this project will provide information on potential pathogenic features of an important aquaculture pathogen and should eventually lead to development of therapeutics to treat, or vaccines to prevent, S. iniae disease. It may also allow development of diagnostic tools for S. iniae. The sequence can also be used for comparative genomics with other sequenced streptococci to identify differences and similarities between human and animal pathogens. Though not a defined goal of this project, an additional intellectual impact that may result is the development or improvement of genome assembly and analysis tools.

Publications

  • No publications reported this period